Applied Microbiology and Biotechnology

, Volume 89, Issue 4, pp 1039–1049 | Cite as

Isolation and characterisation of lactic acid bacterium for effective fermentation of cellobiose into optically pure homo l-(+)-lactic acid

  • Mohamed Ali Abdel-Rahman
  • Yukihiro Tashiro
  • Takeshi Zendo
  • Keisuke Shibata
  • Kenji Sonomoto
Biotechnological Products and Process Engineering


Effective utilisation of cellulosic biomasses for economical lactic acid production requires a microorganism with potential ability to utilise efficiently its major components, glucose and cellobiose. Amongst 631 strains isolated from different environmental samples, strain QU 25 produced high yields of l-(+)-lactic acid of high optical purity from cellobiose. The QU 25 strain was identified as Enterococcus mundtii based on its sugar fermentation pattern and 16S rDNA sequence. The production of lactate by fermentation was optimised for the E. mundtii QU25 strain. The optimal pH and temperature for batch culturing were found to be 7.0°C and 43°C, respectively. E. mundtii QU 25 was able to metabolise a mixture of glucose and cellobiose simultaneously without apparent carbon catabolite repression. Moreover, under the optimised culture conditions, production of optically pure l-lactic acid (99.9%) increased with increasing cellobiose concentrations. This indicates that E. mundtii QU 25 is a potential candidate for effective lactic acid production from cellulosic hydrolysate materials.


l-Lactic acid production Glucose Cellobiose Mixed sugars Enterococcus mundtii 



M.A. Abdel-Rahman is supported by an Egyptian government scholarship offered by the Ministry of Higher Education and Scientific Research, Egypt.


  1. Abe S, Takagi M (1991) Simultaneous saccharification and fermentation of cellulose to lactic acid. Biotechnol Bioeng 37:93–96CrossRefGoogle Scholar
  2. Adnan AFM, Tan IKP (2007) Isolation of lactic acid bacteria from Malaysian foods and assessment of the isolates for industrial potential. Bioresour Technol 98:1380–1985CrossRefGoogle Scholar
  3. Adsul M, Khire J, Bastawde K, Gokhale D (2007) Production of lactic acid from cellobiose and cellotriose by Lactobacillus delbrueckii mutant Uc-3. Appl Environ Microbiol 73:5055–5057CrossRefGoogle Scholar
  4. Anuradha R, Suresh AK, Venkatesh KV (1999) Simultaneous saccharification and fermentation of starch to lactic acid. Process Biochem 35:367–375CrossRefGoogle Scholar
  5. Benthin S, Villadsen J (1995) Production of optically pure d-lactate by Lactobacillus bulgaricus and purification by crystallisation and liquid/liquid extraction. Appl Microbiol Biotechnol 42:826–829CrossRefGoogle Scholar
  6. Bustos G, Moldes AB, Cruz JM, Dominguez JM (2005) Production of lactic acid from vine-trimming wastes and viticulture lees using a simultaneous saccharification fermentation method. J Sci Food Agric 85:466–472CrossRefGoogle Scholar
  7. Caminal G, Lbpez-Santin J, Sola C (1985) Kinetic modeling of the enzymatic hydrolysis of pretreated cellulose. Biotechnol Bioeng 27:1282–1290CrossRefGoogle Scholar
  8. Collins MD, Farrow JAE, Jones D (1986) Enterococcus mundtii sp. nov. Int J Syst Bacteriol 36:8–12CrossRefGoogle Scholar
  9. Ding SF, Tan TW (2006) l-Lactic acid production by Lactobacillus casei fermentation using different fed-batch feeding strategies. Process Biochem 41:1451–1454CrossRefGoogle Scholar
  10. Eberhart BM (1961) Exogenous enzymes of Neurospora conidia and mycelia. J Cell Comp Physiol 58:11–16CrossRefGoogle Scholar
  11. Fujii M, Mori J, Homma T, Taniguchi M (1995) Synergy between an endoglucanase and cellobiohydrolases from Trichoderma koningii. Chem Eng J 59:315–319Google Scholar
  12. Ghosh TK, Das K (1971) Kinetics of hydrolysis of insoluble cellulose by cellulase. Adv Biochem Eng 1:49–55Google Scholar
  13. Gusakov AV, Sinitsyn AP (1992) A theoretical analysis of cellulase product inhibition: effect of cellulase binding constant, enzyme/substrate ratio, and beta-glucosidase activity on the inhibition pattern. Biotechnol Bioeng 40:663–672CrossRefGoogle Scholar
  14. Hofvendahl K, Hahn-Hägerdal B (1997) l-lactic acid production from whole wheat flour hydrolysate using strains of Lactobacilli and Lactococci. Enzyme Microb Technol 20:301–307CrossRefGoogle Scholar
  15. Hofvendahl K, Hahn-Hägerdal B (2000) Factors affecting the fermentative lactic acid production from renewable resources. Enzyme Microb Technol 26:87–107CrossRefGoogle Scholar
  16. Holtzapple M, Cognata M, Shu Y, Hendrickson C (1990) Inhibition of Trichoderma reesei cellulase by sugars and solvents. Biotechnol Bioeng 36:275–287CrossRefGoogle Scholar
  17. Hujanen M, Linko YY (1996) Effect of temperature and various nitrogen sources on l-(+)-lactic acid production by Lactobacillus casei. Appl Microbiol Biotechnol 45:307–313CrossRefGoogle Scholar
  18. John RP, Nampoothiri KM, Pandey A (2006) Simultaneous saccharification and fermentation of cassava bagasse for l-(+)-lactic acid production using Lactobacilli. Appl Biochem Biotechnol 134:263–272CrossRefGoogle Scholar
  19. Joshi DS, Singhvi MS MS, Khire JM, Gokhale DV (2010) Strain improvement of Lactobacillus lactis for d-lactic acid production. Biotechnol Lett 32:1573–6776CrossRefGoogle Scholar
  20. Junco MTT, Martin MG, Toledo MLP, Gomez PL, Barrasa JLM (2001) Identification and antibiotic resistance of fecal enterococci isolated from water samples. Int J Hyg Environ Health 203:363–368CrossRefGoogle Scholar
  21. Kim J-H, Shoemaker SP, Mills DA (2009) Relaxed control of sugar utilization in Lactobacillus brevis. Microbiology 155:1351–1359CrossRefGoogle Scholar
  22. Kim J-H, Block DE, Shoemaker SP, Mills DA (2010) Conversion of rice straw to bio-based chemicals: an integrated process using Lactobacillus brevis. Appl Microbiol Biotechnol 86:1375–1385CrossRefGoogle Scholar
  23. Kruus K, Andreacchi A, Wang WK, Wu JH (1995) Product inhibition of the recombinant CelS, an exoglucanase component of the Clostridium thermocellum cellulosome. Appl Microbiol Biotechnol 44:399–404CrossRefGoogle Scholar
  24. Lee YH, Fan LT (1983) Kinetic studies of enzymatic hydrolysis insoluble cellulose (II). Biotechnol Bioeng 25:959–966Google Scholar
  25. Lunt J (1998) Large-scale production, properties and commercial applications of polylactic acid polymers. Polym Degrad Stab 59:145–152CrossRefGoogle Scholar
  26. Manero A, Blanch AR (1999) Identification of Enterococcus spp. with a biochemical key. Appl Environ Microbiol 65:5173–5176Google Scholar
  27. Medve J, Karlsson J, Lee D, Tjerneld F (1998) Hydrolysis of microcrystalline cellulose by cellobiohydrolase I and endoglucanase II from Trichoderma reesei: adsorption, sugar production pattern, and synergism of the enzymes. Biotechnol Bioeng 59:621–634CrossRefGoogle Scholar
  28. Moldes AB, Alonso JL, Parajo JC (2001) Strategies to improve the bioconversion of processed wood into lactic acid by simultaneous saccharification and fermentation. J Chem Technol Biotechnol 76:279–284CrossRefGoogle Scholar
  29. Nidetzky B, Steiner S, Hayn M, Claeyssens M (1994) Cellulose hydrolysis by the cellulases from Trichoderma reesei: a new model for synergistic interaction. Biochem J 29:705–710Google Scholar
  30. Okano K, Zhang Q, Yoshida S, Tanaka T, Ogino C, Fukuda H, Kondo A (2010) d-lactic acid production from cellooligosaccharides and β-glucan using l-LDH gene-deficient and endoglucanase-secreting Lactobacillus plantarum. Appl Microbiol Biotechnol 85:643–650CrossRefGoogle Scholar
  31. Persson I, Tjerneld F, Hahn-Hägerdahl B (1991) Fungal cellulolytic enzyme production: a review. Process Biochem 26:65–74CrossRefGoogle Scholar
  32. Ramos LP, Saddler JN (1994) Enzyme recycling during fed-batch hydrolysis of cellulose derived from steam-exploded Eucalyptus vitaminalis. Appl Biochem Biotechnol 45:193–207CrossRefGoogle Scholar
  33. Romero-Garcia S, Hernández-Bustos C, Merino E, Gosset G, Martinez A (2009) Homolactic fermentation from glucose and cellobiose using Bacillus subtilis. Microb Cell Fact 8–23. doi: 10.1186/1475-2859-8-23.
  34. Saitoh S, Ishida N, Onishi T, Tokuhiro K, Nagamori E, Kitamoto K, Takahashi H (2005) Genetically engineered wine yeast produces a high concentration of l-lactic acid of extremely high optical purity. Appl Environ Microbiol 71:2789–2792CrossRefGoogle Scholar
  35. Sawa N, Zendo T, Kiyofuji J, Fujita K, Himeno K, Nakayama J, Sonomoto K (2009) Identification and characterization of lactocyclicin Q, a novel cyclic bacteriocin produced by Lactococcus sp. strain QU 12. Appl Environ Microbiol 75:1552–1558CrossRefGoogle Scholar
  36. Severson DK, Barrett CL (1995) Lactobacillus delbrueckii ssp. bulgaricus strain and fermentation process for producing l-(+)-lactic acid. US Patent 5,416,020Google Scholar
  37. Shen X, Xia L (2004) Production and immobilization of cellobiase from Aspergillus niger ZU-07. Process Biochem 39:1363–1367CrossRefGoogle Scholar
  38. Shibata K, Flores DM, Kobayashi G, Sonomoto K (2007) Direct l-lactic acid fermentation with sago starch by a newly-isolated lactic acid bacterium, Enterococcus facieum. Enzyme Microb Technol 41:149–155CrossRefGoogle Scholar
  39. Singhvi M, Joshi D, Adsul M, Varma A, Gokhale D (2010) d-(–)-Lactic acid production from cellobiose and cellulose by Lactobacillus lactis mutant RM2-24. Green Chem 12:1106–1109CrossRefGoogle Scholar
  40. Stockton BC, Mitchell DJ, Grohmann K (1991) Optimum β-glucosidase supplementation of cellulase for efficient conversion of cellulose to glucose. Biotechnol Lett 13:57–62CrossRefGoogle Scholar
  41. Tanaka T, Hoshina M, Tanabe S, Sakai K, Ohtsubo S, Taniguchi M (2006) Production of d-lactic acid from defatted rice bran by simultaneous saccharification and fermentation. Bioresour Technol 97:211–217CrossRefGoogle Scholar
  42. Tokuhiro K, Ishida N, Kondo A, Takahashi H (2008) Lactic fermentation of cellobiose by a yeast strain displaying β-glucosidase on the cell surface. Appl Microbiol Biotechnol 79:481–488CrossRefGoogle Scholar
  43. Tolan JS, Foody B (1999) Cellulases from submerged fermentation. Adv Biochem Eng Biotechnol 65:41–67Google Scholar
  44. Tsai SP, Coleman RD, Moon SH, Schneider KA, Millard CS (1993) Strain screening and development for lactic acid fermentation. Appl Biochem Biotechnol 39(40):323–335CrossRefGoogle Scholar
  45. Venkatesh KV (1997) Simultaneous saccharification and fermentation of cellulose to lactic acid. Bioresour Technol 62:91–98CrossRefGoogle Scholar
  46. Wee YJ, Yun JS, Park DH, Ryu HW (2005) Isolation and characterization of a novel lactic acid bacterium for the production of lactic acid. Biotechnol Bioprocess Eng 10:23–28CrossRefGoogle Scholar
  47. Wee YJ, Kim JN, Ryu HW (2006) Biotechnological production of lactic acid and its recent applications. Food Technol Biotechnol 44:163–172Google Scholar
  48. Xia L, Shen X (2004) High-yield cellulase production by Trichoderma reesei ZU-02 on corn cob residue. Bioresour Technol 91:259–262CrossRefGoogle Scholar
  49. Yáñez R, Moldes AB, Alonso JL, Parajó JC (2003) Production of d-(–)-lactic acid from cellulose by simultaneous saccharification and fermentation using Lactobacillus coryniformis subsp. torquens. Biotechnol Lett 25:1161–1164CrossRefGoogle Scholar
  50. Yoon HH (1997) Simultaneous saccharification and fermentation of cellulose for lactic acid production. Biotechnol Bioprocess Eng 2:101–104CrossRefGoogle Scholar
  51. Yun JS, Wee YJ, Kim JN, Ryu HW (2004) Fermentative production of dl-lactic acid from amylase-treated rice and wheat brans hydrolyzate by a novel lactic acid bacterium Lactobacillus sp. Biotechnol Lett 26:1613–1616CrossRefGoogle Scholar
  52. Zendo T, Eungruttanagorn N, Fujioka S, Tashiro Y, Nomura K, Sera Y, Kobayashi G, Nakayama J, Ishizaki A, Sonomoto K (2005) Identification and production of a bacteriocin from Enterococcus mundtii QU 2 isolated from soybean. J Appl Microbiol 99:1181–1190CrossRefGoogle Scholar
  53. Zhang B, He PJ, Ye NF, Shao LM (2008) Enhanced isomer purity of lactic acid from the non-sterile fermentation of kitchen wastes. Bioresour Technol 99:855–862CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Mohamed Ali Abdel-Rahman
    • 1
    • 2
  • Yukihiro Tashiro
    • 3
  • Takeshi Zendo
    • 1
  • Keisuke Shibata
    • 1
  • Kenji Sonomoto
    • 1
    • 4
  1. 1.Laboratory of Microbial Technology, Division of Applied Molecular Microbiology and Biomass Chemistry, Department of Bioscience and Biotechnology, Faculty of Agriculture, Graduate SchoolKyushu UniversityHigashi-kuJapan
  2. 2.Botany and Microbiology Department, Faculty of Science (Boys)Al-Azhar UniversityCairoEgypt
  3. 3.Department of Life StudySeinan Jo Gakuin University Junior CollegeKitakyushu CityJapan
  4. 4.Laboratory of Functional Food Design, Department of Functional Metabolic Design, Bio-Architecture CenterKyushu UniversityHigashi-kuJapan

Personalised recommendations